Nucleic acids and vectors encoding human defensin polypeptide and applications thereof

Abstract

The present invention relates to a novel human polypeptide defensin Def-X, genomic DNA and cDNA encoding Def-X, vectors containing Def-X encoding polynucleotides, and cells transformed with these vectors. The invention also provides for the use of Def-X polypeptides as an antibiotic, a cytotoxic, repair, and endocrine regulatory agent, and as pesticide. Def-X can also be used as agent, a cosmetic or pharmaceutical compositions for the treatment of microbial infections, in particular bacterial, fungal, and viral infections, parasitic infections, cancer, inflammation, and immune deficiencies. The invention provides diagnostic methods and kits for the determination of a microbial or parasitic infection and/or inflammation; also provided are methods of screening for predisposition to immune deficiencies or cancer.

1. An isolated polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 3; or 
 b) a polypeptide consisting ofa fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of SEQ ID NO: 3; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       2. The isolated polynucleotide according to  claim 1 , wherein said polynucleotide encodes a polypeptide consisting ofa fragment of SEQ ID NO: 3 comprising at least 15 consecutive amino acids of the polypeptide of SEQ ID NO: 3 wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       3. The isolated polynucleotide according to  claim 1 , wherein said polynucleotide encodes a polypeptide comprising the sequence of SEQ ID NO: 3. 
     
     
       4. The isolated polynucleotide according to  claim 1 , wherein said polynucleotide encodes a polypeptide consisting of a fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of the polypeptide of SEQ ID NO: 3 wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       5. The isolated polynucleotide according to  claim 1 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a Fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of SEQ ID NO: 3 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       6. The isolated polynucleotide according to  claim 2 , wherein said polynucleotide encodes a polypeptide comprising a mature peptide consisting of the sequence of SEQ ID NO: 6. 
     
     
       7. An isolated polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 6; and 
 b) a polypeptide consisting of a fragment comprising at least 10 consecutive amino acids of the sequence of SEQ ID NO: 6; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       8. The isolated polynucleotide according to  claim 7 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of SEQ ID NO: 6 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       9. The isolated polynucleotide according to  claim 7 , wherein said polynucleotide encodes a polypeptide comprising the sequence of SEQ ID NO: 6. 
     
     
       10. The isolated polynucleotide according to  claim 7 , wherein said polynucleotide encodes a polypeptide consisting of a fragment comprising at least 10 consecutive amino acids of the sequence of SEQ ID NO: 6 wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       11. The isolated polynucleotide according to  claim 7 , wherein said polynucleotide encodes a polypeptide consisting of a fragment comprising at least 15 consecutive amino acids of the sequence of SEQ ID NO: 6 wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       12. A vector comprising a polynucleotide encoding a polypeptide selected form the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 3; and 
 b) a polypeptide consisting of a fragment comprising at least 10 consecutive amino acids of SEQ ID NO: 3; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       13. The vector according to  claim 12 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of SEQ ID NO: 3 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       14. The vector according to  claim 12 , further comprising elements ensuring the expression of said polynucleotide in a host cell. 
     
     
       15. A vector comprising a polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 6; and 
 b) a polypeptide consisting of fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of the sequence of SEQ ID NO: 6; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       16. The vector according to  claim 15 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of SEQ ID NO: 6 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       17. The vector according to  claim 15 , further comprising elements ensuring the expression of said polynucleotide in a host cell. 
     
     
       18. A host cell transformed with a vector comprising a polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypetide comprising the sequence of SEQ ID NO: 3; and 
 b) a polypeptide consisting of a fragment comprising at least 10 consecutive amino acids of SEQ ID NO: 3; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity the group consisting of antimicrobial activity and cytotoxic activity. 
 
     
     
       19. The host cell according to  claim 18 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of SEQ ID NO: 3 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       20. The host cell according to  claim 18 , wherein said vector further comprises elements ensuring the expression of said polynucleotide in said host cell. 
     
     
       21. A host cell comprising a vector comprising a polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 6; and 
 b) a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of the sequence of SEQ ID NO; 6; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       22. The host cell according to  claim 21 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of SEQ ID NO: 6 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       23. The host cell according to  claim 21 , wherein said vector further comprises elements ensuring the expression of said polynucleotide in said host cell. 
     
     
       24. A method of producing a polypeptide comprising culturing a host cell transformed with a vector comprising a polynucleotide encoding a polypeptide selected from the group consisting or:
 a) a polypeptide comprising the sequence of SEQ ID NO: 3; and 
 b) a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of SEQ ID NO: 3; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       25. The method according to  claim 24 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 3 comprising at least 10 consecutive amino acids of SEQ ID NO: 3 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       26. The method according to  claim 24 , wherein said vector further comprises elements ensuring the expression of said polynucleotide in said host cell. 
     
     
       27. A method of producing a polypeptide comprising culturing a host cell transformed with a vector comprising a polynucleotide encoding a polypeptide selected from the group consisting of:
 a) a polypeptide comprising the sequence of SEQ ID NO: 6; and 
 b) a polypeptide consisting of a fragment of SEQ ID NO: 6 of at least 10 consecutive amino acids of the sequence of SEQ ID NO: 6; wherein said fragment has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
 
     
     
       28. The method according to  claim 27 , wherein said polynucleotide comprises the sequence of SEQ ID NO:2 or a fragment thereof wherein said fragment encodes a polypeptide consisting of a fragment of SEQ ID NO: 6 comprising at least 10 consecutive amino acids of SEQ ID NO: 6 and has at least one biological activity selected from antimicrobial activity or cytotoxic activity. 
     
     
       29. The method according to  claim 27 , wherein said vector further comprises elements ensuring the expression of said polynucleotide in said host cell.